The protein glycosylation pathway still holds many questions in the case of lower eukaryotes. Thus, while the characteristics of some carbohydrate chains isolated from glycoproteins of certain filamentous fungi such as Aspergillus and Trichoderma resemble those found in yeast, others resemble those encountered in mammals. In an attempt to shed some light on the glycosylation in filamentous fungi, we have studied the predominant N-glycans from cellobiohydrolase 1, secreted from Trichoderma reesei RUTC 30. One of the compounds found is a novel phosphate-containing oligosaccharide that, to the best of our knowledge, has not been reported before. The primary structure of the novel phosphate-containing oligosaccharide, isolated from T reesei cellobiohydrolase 1, was determined by ES1 MS and found to comprise Glcl-6P03-Man7-9GlcNAC2. The Glu extensions appear to be on the lower 3-branched antenna. Thepresence of mannose residues linked through a phosphate diester resembles glycosyl synthesisin yeast. Several additional samples are under investigation.